Project Summary/Abstract All tissues of a living organism rely on normal thyroid hormone levels to develop and function properly. Millions of people world-wide suffer from thyroid dysfunction (TD). TD incidence is 4-10% in the general population and is considerably higher in people with metabolic diseases. Despite the central role that thyroid gland plays in human physiology, most of what we know about thyroid cell biology, control of thyroid follicular cell growth, vasculature adaptation and endocrine interactions has been derived from histological assessments and in vitro studies that limit translation to the in vivo environment. Remarkably, these in vitro set-ups do not replicate the basic functional unit of the thyroid, particularly the three-dimensional angio-follicular unit. Therefore, in vivo studies of thyroid gland physiology and pathophysiology are hindered by the lack of approaches in which the thyroid gland can be assessed in its natural architecture. The long-term goal of my research is to understand the cell biology of the thyroid gland in the living organism. The strength of the present application relies on my preliminary data of a robust technological platform that I developed in my laboratory. This novel approach allows noninvasive real-time in vivo imaging of the mouse and human follicular cell. The technology is based on transplantation of the mouse and human thyroid fragments into the mouse eye, which engrafts and becomes revascularized; it permits the analysis of longitudinal changes in thyroid graft size, blood flow and vessel diameters, with three-dimensional in vivo images of the thyroid angio-follicular unit. My hypothesis is that mouse and human thyroid gland transplanted into the mouse anterior chamber of the eye resembles the normal cellular structure of the thyroid and recapitulate the responses/adaptations to iodine deficiency and thyrotoxicosis. To test my hypothesis, I will pursue two specific aims: 1) obtain unique in vivo intracellular physiological measurements of the angio-follicular unit and its response to physiological stimuli and 2) gain in vivo insights into the adaptation of the human and mouse angio-follicular unit and its cellular components to iodine deficiency and thyrotoxicosis. I will study mice transplanted with mouse and human thyroid specimens and manipulate different endocrine stimuli. Transgenic mouse models will allow me to demonstrate for the first time thyrocytes activation in vivo. I will challenge the system by hyperactivating the thyroid gland with experimental iodine deficiency. The responses to thyrotoxicosis (increased levels of thyroid hormone) will also be assessed. This innovative platform will overcome a major technical roadblock by allowing the visualization and measurement of dynamic properties of the thyroid gland biology in vivo. At their successful completion, my studies will enhance our knowledge of the in vivo mechanisms of vasculature adaptation and thyroid follicular cell growth. I predict that deploying these methods will provide unprecedented opportunities to advance in the understanding of human thyroid physiology and thyroid malignancies.